This application stems from our recent findings that i) anti-human high molecular weight-melanoma associated antigen (HMW-MAA) immunity elicited by mouse anti-id mAb MK2-23 in about 60% of patients with advanced melanoma is associated with regression of lesions in a few patients and with a statistically significant survival prolongation; ii) major histocompatibility complex plays a role in the immune response of mice to anti-id mAb MK2-23 and iii) low doses of systemic IL-2 markedly enhance the immunogenicity of mAb MK2-23 in patients with melanoma. Therefore, we plan to continue our studies to define the factors regulating the immune response to anti-id mAb MK2-23 and to enhance the immunogenicity of anti- id mAb. Specifically, we will continue to investigate the effect of the disease stage and the role of the HLA region in the development of anti- HMW-MAA immune response. The resulting information may aid to select patients who may benefit from immunotherapy with anti-id mAb. Furthermore we will explore in animal model systems approaches to enhance anti-HMW-MAA immune response by testing i) new adjuvants, ii) combinations of anti-id mAb which bear the internal image of distinct determinants of HMW-MAA and iii) fusion proteins between anti-id mAb and IL-2 and by defining the effect of antibodies to constant regions of Ig on the immunogenicity of anti-id mAb. Recurrence of melanoma in immunized patients in spite of anti-HMW-MAA immune response is likely to reflect, at least in part, low or lack of HMW-MAA expression in melanoma lesions. Therefore, this proposal aims at testing the hypothesis that the efficacy of active specific immunotherapy may be improved in patients with melanoma by combining anti-id mAb which bear the internal image of distinct determinants of HMW-MAA and of melanoma associated antigens with a non coordinate expression in melanoma lesions. To this end the immunogenicity and toxicity of the combination of anti-id mAb with this specificity will be tested in rabbits, which express both HMW-MAA and GD/3 ganglioside in their normal tissues. Rabbit immune sera will be tested for their ability to mediate immune lysis of human melanoma cells with differential expression of HMW-MAA and GD/3 ganglioside. The information resulting from studies in animal model systems will be utilized to design the immunization schedule with the most effective adjuvant and combination of anti-id mAb in patients with advanced melanoma in a phase I-II clinical trial.